We investigated the possibility of selective in vivo transduction of growing brain tumors with retroviral vectors carrying the herpes simplex thymidine kinase (HS-tk) gene to conger drug sensitivity to the antiviral drug ganciclovir (GCV). We have shown that rats with a malignant brain tumor which were given an intratumoral stereotaxic injection of murine fibroblasts that were producing the HS-tk vector and then treated with GCV had regression of their tumors. Rats treated with a control vector producer cell line containing the beta-galactosidase gene (lacZ gene) developed large tumors and died. Using the lacZ gene as a reporter gene, we were able to study the dynamics of in situ tumor transduction in rats. Dose-response studies of GCV and various concentrations of the injected vector-producer cells provided further data to guide us in designing a clinical protocol for the treatment of patients with brain tumors. No significant toxicity was observed in toxicity studies in mice, rats, and nonhuman primates. Long-term survival experiments using this approach showed a cure rate of 30-35% and significant prolongation of survival in the remainder of the treated rats. Survival curves suggest the need for repeat treatment to enhance the efficacy of our approach. Based upon these findings, we have proposed a human clinical trial to determine trial to determine the efficacy of HS-tk transduction in patients with brain tumors. Retroviral-mediated gene transfer into experimental brain tumors using cytokine genes (IL-2), alone or as a combined HS-tk/IL-2 vector, were also evaluated. However, no enhancement of tumor eradication was observed with the addition of the cytokine gene confirming the limited usefulness of immune enhancement within the CNS.